We seek NIH funding to develop a vaccine against cryptosporidiosis for children in LMIC. As no effective therapies exist, there is a critical need for vaccination to reduce morbidity and mortality attributed to cryptosporidiosis, which is estimated to cause diarrhea in >10 million children and kills more than half a million children each year. The Global Enteric Multicenter Study (GEMS) ranked Cryptosporidium as the 2nd most prevalent pathogen (1st rotavirus; 3rd Enterotoxigenic E. coli; 4th Shigella spp.) causing moderate-to-severe diarrhea (MSD) in children in 4 sites in sub-Saharan Africa and 3 in South Asia. The GEMS further made the case that controlling these 4 pathogens will eliminate > 90% of MSD in children. Much progress had been made towards this goal in that vaccines already exist for rotavirus or are in clinical trials for ETEC and Shigella. However, vaccine development and testing against Cryptosporidium, a eukaryote with many serious technical obstacles, lags, and there has been minimal scientific progress to overcome these challenges since the first few surface proteins were discovered more than 20 years ago. To address these gaps, we identified novel parasite antigens, developed novel immunocompetent rodent models of infection, and deployed innovative intradermal mRNA immunization technology developed by our CureVac partners. In this proposal, we demonstrate the experience, skills, innovative mRNA vaccine technology and use of rodent models and reagents to identify an effective panel of mRNAs against cryptosporidiosis as the first step. Using reverse vaccinology, immunoproteomics, gene expression, and comparative genomics to other related protozoa, we identified a panel of 32 novel candidates derived from C. hominis and used by CureVac to generate mRNAs. Specific Aim 1 focuses on screening 32 (or more as needed) mRNAs encoding C. hominis in the enteric mouse model intradermally and orally challenged with C. tyzzeri to identify a panel (~4-6) immunogens, which will then be confirmed in the IT rat model which is susceptible to infections with C. hominis or C. parvum. Specific Aim 2 will identify immune mechanisms that drive vaccine-induced protection in the two rodent models. This proposal, if awarded, will lead to identifying a panel of immunogens that together protect mice against challenge with C. tyzzeri; and rat trachea against infections with C. hominis and C. parvum. We anticipate that the proposed research will form the first step leading ultimately to the development of an effective vaccine to protect infants and children at risk, mostly in LMIC.